B 5:0-3% 5:9-25% 5:51-75% Introduction The Berkeley Drosophila Transcription Network Project (BDTNP) is a multidisciplinary collaboration studying the developmental regulatory network of Drosophila blastoderm embryos. One component of this project (Luengo et al., Genome Biology 7:R123, 2006) maps the full 3D blastoderm expression patterns of 37 principal developmental regulatory genes and hundreds of their targets at cellular resolution, and uses these data to model potential regulatory interactions. We have developed an automated pipeline and methods for producing these data. Both the algorithms and the data are freely available at: C.L. Luengo Hendriks 1, C.C. Fowlkes 2, S.V.E. Keränen 1, L. Simirenko 1, G.H. Weber 3, O. Rübel 4, M.-Y. Huang 4, A.H. DePace 1, C. Henriquez 1, X.-Y. Li 1, H.C. Chu 1, D.W. Kaszuba 1, A. Beaton 1, S. Celniker 1, B. Hamann 4, M.B. Eisen 1, J. Malik 2, D.W. Knowles 1 and M.DBiggin 1 Virtual Embryos As Tools For 3D Gene Expression Analyses Uses of Virtual Embryos The use of standardized virtual embryos allows temporal comparison within each nucleus between earlier expression of regulators in one cohort and the later expression of target gene patterns in another cohort, as well as better estimates of the developmental increase in complexity. 1) Life Sciences and Genomics Division, Lawrence Berkeley National Laboratory 2) Computer Science Division, University of California, Berkeley 3) Computational Research Division, Lawrence Berkeley National Laboratory 4) Institute for Data Analysis and Visualization, University of California, Davis Generating Virtual Embryos Because each imaged embryo contains expression information of only two genes, expression data from hundreds of embryos are mapped onto a virtual embryo to allow many genes’ expression to be compared and modeled within each cohort. These virtual embryos contain nuclei placed to match the average density pattern and embryo shape for each cohort. PointClouds Virtual Embryo co-stainedcoarse registration fine registrationaverages eve gt eve gt eve gt eve gt co-stained coarse fine Density changes through time (Keränen et al., Genome Biology 7:R124, 2006) (Fowlkes et al. in preparation) For example, the spatial expression of D, Kr, gt and slp1 change during stage 5. In virtual embryos, such changes in multiple genes can be computationally analyzed in a standardized environment. The expression of eve in cohort 6 (stage 5: %) (A, B) compared to the expression patterns of gap genes in cohort 4 (stage 5:26-50%) (C, D). The expression levels of each eve stripe can be visualized and analyzed separately (B), and the relative expression levels of their putative regulators can be then studied in the corresponding nuclei either by analyzing their clustering in 3D scatter plots (C) or by observing the multigene expression profiles in parallel co-ordinate view (D) that also can contain 1D spatial information. A C B D Mutants & Other Species Gene expression data in regulatory factor mutant embryos and other Drosophila species is also being collected. Unrolled view on comparison of gt mRNA expression in D. melanogaster (green) and D. pseudoobscura (red) aligned on eve mRNA expression (below). Note the relative positions of anterior gt stripes. Dorsal-ventral signals affect the anterior-posterior pattern formation (Keränen et al., Genome Biology 7:R124, 2006). In dorsalized gd 7 mutants the ventral ftz stripes resemble wild type dorsal ftz stripes (A), whereas in ventralized Toll 10B mutants the dorsal ftz stripe positions resemble the wild type ventral ftz stripes (B). The intensity profiles of the stripes lose the dorsoventral polarity (D, F) that is seen in wild type looking embryos (C, E). CD EF □ wild type □ mutant Collecting the Data The 3D, full blastoderm images are analyzed and accurately converted into PointCloud files by an automated algorithm. We currently have data for 24 of the principal regulators and over 80 putative target genes, the latter selected using BDTNP ChIP-chip binding data and BDGP expression data. 5:0-3% 5:4-8% 5:9-25% 5:26-50% 5:51-75% 5:76-100% D Dfd Kr bcd brk cad croc eve fkh ftz gt h hb hkb kni knrl